1. Field of the Invention
The present invention relates generally to improved methods for tissue engineering including tissue transplantation, augmentation and regeneration. More particularly, the present invention provides a method for the generation of donor vascularized tissue suitable for use in tissue transplantation, augmentation and/or repair. The present invention further enables the use of a support matrix in the generation of an anatomical construct comprising the donor vascular tissue. The support matrix may be devised such that it has dimensions of a size and shape adapted to simulate those of tissue to be transplanted, augmented and/or repaired. In addition to its use in tissue repair, the methods and support matrix of the present invention may also find application as a means for delivering a desirable gene product to a subject. The method and support matrix of the present invention is conveniently be made available in the form of a kit, for use generally in the field of tissue engineering.
2. Description of the Related Art
Bibliographic details of the publications referred to in this specification are also collected at the end of the description.
Reference to any prior art in this specification is not, and should not be taken as, an acknowledgment or any form of suggestion that this prior art forms part of the common general knowledge in any country.
Tissue engineering utilizing homologous starting material offers the prospect of replacing missing or non-functioning body parts with newly created, living tissue. It has the potential to minimize loss of tissue and resultant pain from the donor site experienced in conventional reconstructive surgery or to recreate specialized tissue for which there is no donor site, while obviating the long-term immunosuppression required for heterologous transplantation.
It combines the techniques of tissue culture, the creation of bio-compatible materials and the manipulation of angiogenesis in order to create new, vascularized tissue to replace damaged tissue or tissue which is congenitally absent.
One of the major challenges faced in tissue engineering is to create differentiated tissue of the appropriate size and shape. Tissue created without a functional vasculature is strictly limited in size by the constraints of oxygen diffusion; if the tissue is too large it will become necrotic before the host has time to create a new blood vessel supply. Thus there are many advantages in creating new tissue containing a functional vasculature. Additionally, as the new tissue may need to be produced at a site on the body remote from the defect, or on an immuno-suppressed carrier animal or in vitro with an extra-corporeal circulation, the blood supply for the new tissue must be defined, so that it can be brought with the tissue intact to the site of reconstruction.
The creation of skin flaps, a living composite of skin and its underlying fat, is a common technique used to repair tissue defects in reconstructive surgery. Because these flaps must retain their blood supply to remain viable after transplantation, the origin of the flaps is limited to those areas where there is an anatomically recognized blood vessel source. In order to overcome this limitation, skin flaps can be “pre-fabricated” by implanting short segments of blood vessels into a desired site, and utilizing the resultant angiogenesis to vascularize a flap of the desired size and composition. Subsequently this vascularized flap can be transferred by microsurgery to the region of interest. This technique is, however, limited by the availability of donor tissue, and the disfigurement that results at the donor site.
In an extension to this technique, Erol and Spira, Surgery 66: 109-115, 1980 demonstrated that the creation of an anastomosed arteriovenous (AV) loop beneath a skin graft could produce a vascularized skin “flap”.
However, while the generation of vascularized skin using an AV loop has been demonstrated, the production of other vascularized tissues suitable for grafting remains elusive. Vascularized adipose tissue, for example, is often demanded in reconstructive procedures; however, donor mature adipose tissue is extremely fragile, and will rapidly become necrotic if not immediately reconnected to a functional blood supply. Furthermore, the use of conventional autologous transplantation techniques involves “robbing Peter to pay Paul”, producing disfigurement at the donor site. The ability to produce new tissue with a defined vasculature would overcome this major shortcoming.
Khouri et al., Surgery 114: 374-380, 1993 and Tanaka et al., Jpn. PRS 16: 679-686, 1996 have demonstrated that an arteriovenous loop could intrinsically generate new, vascularized tissue when it was lifted from the body, sandwiched between sheets of collagenous matrix and isolated from the surrounding tissue within a plastic chamber. In the model described by Khouri et al., 1993, supra the generation of new tissue relied on the addition of recombinant BB-homodimer of Platelet-Derived Growth Factor (BB-PDGF), and even with this supplement the tissue was labile, peaking in volume at 15 days and subsiding by 30 days. Similarly, tissue growth in Tanaka's model, where the chamber was supplemented with β-Fibroblast Growth Factor (β-FGF or FGF-2), continued to increase in volume, peaking at two weeks but returned to the levels of the unsupplemented control chambers after four weeks. This AV loop model is not generally known in the field of tissue engineering.
International Patent Application No. PCT/AU01/01031 (International Patent Publication No. WO 02/15914) describes inter alia the use of an AV loop in a fabricated chamber which is implanted into a subject. Tissue was found to successfully grow around the AV loop in a shape dependent on the constraints of the chamber.
Despite the success of the methodology and chamber described in International Patent Publication No. WO 02/15914, there is a need to further improve the growth of tissue around a functional circulatory system.